1. Field of the Invention
The present invention relates to Kombic Acid derivatives as anti-cancer agents. More particularly, the present invention relates to kombic acid bis-succinate (also known as sarganol bis-succinate) and the corresponding alkyl and aryl derivatives as apoptogens with specificity for inducing apoptosis in cancer cells, both in vitro and in vivo. The present invention further relates to a method of synthesizing bis-succinate derivatives of sarganol, particularly for use as an anti-cancer agent.
2. General Background of the Invention
Kombic Acid was first isolated and characterized in 1983 (1). Subsequent work has shown that kombic acid, previously thought to have the structure shown in FIG. 1, was actually identical to several other compounds isolated from different sources and having different names (2). As a result, the chemical structure 12-(2,5-dihydroxy-3-methylphenyl)-6,10-dimethyl-2-(4-methyl-3-pentenyl)-2E,6E,10E)-2,6,10-dodecatrienoic acid has at least 3 common names, all of which refer to the structure shown in FIG. 2; Sarganol, Sargahydroquinoic Acid, and Kombic Acid. For the purposes of clarity, the name Sarganol will be used throughout this application.
Sarganol was isolated form the seedfat of a West African (Ghana, for example) tree called Pycnanthus Angolensis, also termed Pycnanthus Kombo. The seedfat, a vegetable butter, has become known as kombo butter. Two U.S. Patents have been obtained relating to the use of kombic acid in the treatment of coronary artery disease and neoplastic diseases. (3, 4) These patents disclose the antioxidative properties of kombic acid and show the superior antioxidative properties of kombic acid when compared to tocopherols (Vitamin E); generically, a molecule structurally similar to sarganol.
Fats and oils are water-insoluble, hydrophobic substances of vegetable, land animal or marine animal origin that consist mostly of glyceryl esters of fatty acids, called triglycerides. Their structure is shown below, where R1, R2, and R3 can be the same or different —(CH2)xCH3 chains, with x being an even number of 4 or greater.
The chains (the Rs) may be completely saturated with respect to hydrogen or have one or more double bonds. When R1 is seventeen carbons with no double bonds, the chain is called stearic; in this case there are thirty-five hydrogen atoms attached to the seventeen carbons. With one double bond the same carbon-length chain is called oleic and there are thirty-three attached hydrogen atoms. When there is more than one double bond, the fatty acids are polyunsaturated. Linoleic acid, for example, has eighteen carbons atoms and two double bonds, and linolenic acid has three double bonds and eighteen carbon atoms.
There are, annually, roughly 100 million metric tons of fats and oils consumed globally with about 80% used for human food. The balance is used as industrial oils; in animal feed; to make soap; and to produce oleochemicals, which have many industrial applications, most notably as plastics additives and food processing ingredients.
The principal fats and oils used in food are canola, soybean, palm, sunflower seed, coconut, palm kernel, sesame, olive, corn, cottonseed, edible tallow and lard. The most frequently occurring fatty acids found in these fats and oils are stearic (C18:0), oleic (C18:1), linoleic (C18:1), linolenic (C18:3), palmitic (C16:0), myristic (C14:0) and lauric (C12:0). The first two digits in the carbon subscript refer to carbon chain length, and the number after the colon refers to the number of double bonds in the chain.
Myristoleic acid, which has fourteen carbon atoms and one double bond in the chain has an ester derivative, cetyl myristoleate, with claimed efficacy in relieving the pain of rheumatoid arthritis and osteoarthritis1,2. The myristoleic acid used to make this product up to now has been derived from beef tallow.
There are two commonly accepted reference sources with respect to published treatises on fats and oils: “Bailey's Industrial Oil and Fat Products” and Gunstone and Padley's, “Lipid Technologies and Applications.” In “Bailey's” there is the following discussion of sources of myristoleic acid, all based on land animal or marine animal origin.                9-Tetradecenoic (myristoleic) acid is the most common of the tetradecenoic acids, being first detected in whale oil in 1925 at about 1.4%, later in whale blubber oil, in shark liver oil, Antarctic whale oil, eel oil, and turtle oil. In 1924 it was suggested that myristoleic acid occurs in butterfat; it was found to constitute 1% of the total acids. It also occurs in goat milk fat, human milk fat, and various animal depot fats, (particularly beef tallow).It is noteworthy that there is not the slightest reference in Bailey's to any vegetable oil sources.        
Gunstone and Padley, in their well-recognized reference work mention hundreds of fatty acids but make no reference of any sort to myristoleic acid. Useful products can be obtained from myristoleic acid, most notably cetyl myristoleate, a possible remedy for alleviating the pain and inflammation of arthritis and related maladies1,2. However, cetyl myristoleate based on myristoleic acid sourced from animal origins, up to now the only ostensible source, has several disadvantages:                1) Fatty acids derived from beef tallow run the risk, albeit slight, of inducing bovine spongiform encephalitis (mad-cow disease).        2) Any fatty acid sourced from land animal or marine animal origins cannot be Kosher or the Islamic equivalent, Halal.        3) Any fatty acids sourced from land animal or marine animal origins cannot be “vegetarian” or “vegetable-oil food-grade.”        4) Fatty acids sourced from land animal and marine animal origins are identifiable by at least certain animals as having an undesirable taste and/or smell which negatively affects the ability of these animals to orally ingest supplements or drugs made with them.        
Myristoleic acid, however, is not exclusively sourced from non-vegetable oil origins. There is a tree that produces a nut containing a vegetable butter that is a relatively good source of myristoleic acid3,4,5,6. The fat is known as kombo butter. It comes from the seeds of Pycnanthus Kombo (Myristicaceae family) found in West Central Africa. Other compounds isolated from P. Kombo (P. Angolensis) include 2′-hydroxy-4′, 7-dimethoxy isoflavone and 2′-hydroxy fomonometin8. In addition, U.S. Pat. No. 5,674,900 00 describes the isolation and use of terpenoid quinones from the stems and leaves (not the seedfat) of P. kombo for use in treating diabetes9.
The seedfat of P. kombo is reddish-brown and has a distinct aromatic odor. The fat also contains 20-30% of kombic acid. Sarganol is not a fatty acid per se, rather it is a substituted fatty acid, and must be separated and removed from kombo butter in manufacturing downstream oleochemical products such as myristoleic acid. From kombo butter, the unit operations to obtain relatively pure distilled fatty acid mixtures containing appreciable levels of myristoleic acid include: 1) fat (crude kombo butter) saponification to split the fat and form the sodium soaps of the fatty acids, thereby separating and removing the glycerine, 2) acidulation of the sodium soaps of the fatty acids to form the free fatty acids, and 3) molecular distillation of the crude fatty acids for purposes of purification. The cetyl esters can then be formed by conventional esterification reactions. The present invention describes bis-succinate derivatives of sarganol isolated from the seed fat of Pycnanthus Kombo and their use as potent antineoplastic agents with selectivity for cancer cells.